Xenograft Tumour Model

The inhibitory effect of metformin on GC in vivo was determined using a mouse xenograft tumour model. All experiments involving animals were conducted in accordance with the operation procedures approved by the Institutional Committee of Nanjing Medical University for Animal Research. Male nude mice (5–6 weeks old) were randomly divided into 2 groups, with 5 mice per group. The first group received subcutaneous injections into the dorsal flank with 100 µL PBS containing 8.0 × 10⁶ MGC803 cells and daily injections of normal saline (100 μL). The second group received MGC803 cell injections and daily metformin (300 mg/kg) treatment. The metformin treatment started on the third day after inoculation of the GC cells and lasted for 4 weeks. Digital callipers were used to measure the growth of the tumour every three days, and the formula (volume = 0.5 × width² × length) was used to calculate the tumour volume.